This grant proposes a series of related studies on the synthesis and processing of the neuronal trophic factor Nerve Growth Factor (NGF) in sites that are important for neuronal development. Using a cDNA probe encoding the mouse beta-NGF gene, studies will be continued on identifying tissues that synthesize NGF in the adult and during development. We will attempt to identify the cell types that synthesize NGF within these tissues by examining NGF gene expression in defined cell types in culture and by doing in situ hybridization on tissue sections using an NGF gene probe. The NGF probe will be used to quantify NGF mRNA levels during the development of tissues innervated by sympathetic and sensory neurons. We will determine whether there are relationships between the synthesis of NGF mRNA and the induction of sumpathetic neurons, the development and extent of innervation in targets, and neuronal cell death. We will determine in adult tissues whether denervation, already known to increase NGF levels, does so by increasing beta-NGF gene expression. In a second set of experiments, we will examine the form of NGF synthesized in tissues and cells that express the gene using antibodies that are specific for amino acid segments in the precursor. Residues between these segments are cleaved during normal processing in exocrine cells in the male mouse submaxillary gland. We will see whether the same processing steps occur in the tissues and cells believed to be the physiological sources of NGF. Some of these cells are known not to process other prohormones, so the precursor may be important. It will be purified and its properties will be examined. If NGF is processed in physiological sources, we will determine whether proteases are able to process the prohormone in vitro are present at appropriate sites in vivo. Antibodies specific for amino acid residues in the precursor will be used to purify the three polypeptides produced in addition to NGF by processing of the prohormone in the mouse submaxillary gland. These purified polypeptides will be tested for biological actions on NGF-responsive cells and will be used to make antibodies for studies on processing of proNGF. These results should provide information on how NGF, an important trophic hormone, controls the development of responsive neuronal cell populations.